PROJECT SUMMARY: PROJECT 2 Type 1 VWD is characterized by a partial quantitative deficiency of VWF. Although it accounts for 70% of all cases of VWD, the molecular mechanisms underlying the pathogenesis of type 1 VWD remain poorly understood. However, linkage studies have shown that reduced plasma VWF levels (particularly in patients with mild to moderate reductions in the range 30 - 50 IU/dL) are independent of the VWF gene on chromosome 12. Accumulating data from both animal and human studies have demonstrated that the complex glycan determinants expressed on VWF play critical roles in regulating many aspects of its biology, including synthesis, proteolysis and clearance. Aim 1 of this project will investigate how VWF N- and O-glycan structures vary in a large cohort of patients with comprehensively phenotyped VWD already enrolled through 1) the Zimmerman program, (2) the Canadian type 1 VWD study and (3) the Low VWF Ireland Cohort study. Given the complex heterogeneous nature of the N- and O- carbohydrate structures expressed on human VWF, this unique combined clinical resource will for the first time provide adequate power to enable elucidation of the role of VWF glycans in VWD pathogenesis. In Aim 2, a subset of patients with type 1 VWD and Low VWF in whom glycan abnormalities have been identified will be selected for further analysis. In this cohort, the N- and O-glycan structures of VWF will be definitively sequenced using state-of-the-art mass spectroscopy glycoproteomic analysis. Finally, using a number of novel parallel strategies, Aim 3 of the project will focus on (1) investigating the mechanisms underlying abnormal VWF glycosylation in patients with type 1 VWD and Low VWF, and (2) determining whether VWF carbohydrate modifications play a direct causal role in mediating VWD pathogenesis (e.g. by reducing biosynthesis or modulating enhanced clearance).